Histamine signaling in the hypothalamus controls energy expenditure, thermoregulation, sleep and feeding. Transgenic animals in which histamine signaling is disrupted (the H1 receptor k.o., the H3 receptor k.o. and the histamine decarboxylaze k.o.) develop leptin-resistant obesity, indicating the importance of normal histamine signaling for the maintenance of energy homeostasis. Furthermore, leptin-deficient (ob/ob) and leptin receptor- defective (db/db) obese mice display lowered hypothalamic histamine concentrations. These observations underline the key role played by histamine downstream of leptin. Pharmacological agents that increase the hypothalamic concentration of histamine are beneficial in animal models of diabetes, obesity and narcolepsy, and several such drugs are in clinical trials. However, the cellular mechanisms activated by increased histamine signaling in the hypothalamus are not well understood. Previous studies have shown that histamine effects on thermoregulation and energy expenditure are due to its actions in the preoptic area/ anterior hypothalamus (PO/AH). Electrophysiological studies of PO/AH have revealed the existence of thermosensitive neurons that are now considered to be key elements of the thermoregulatory system of the CNS. Warm- sensitive PO/AH neurons increase their firing rate in response to temperature elevation (in contrast to the temperature-insensitive neurons) and function as integrators of homeostatic conditions. PO/AH thermoregulatory neurons project to the dorsomedial hypothalamus or to the rostral raphe pallidus. We will identify thermoregulatory PO/AH neurons by retrograde labeling from these centers. Our primary hypothesis is that histamine affects the activity of PO/AH neurons involved in thermoregulation and we predict that this action determines the changes in body temperature and energy expenditure induced by histamine. We also hypothesize that histamine signaling in the PO/AH is altered in leptin-deficient (ob/ob) mice, a well-known obesity model. Preliminary data indicate that H1, H2 and H3 receptors are present in this region, that histamine potently modulates synaptic activity and postsynaptic conductances in PO/AH neurons and also exerts complex actions on intracellular Ca concentrations. We plan to identify both warm-sensitive and - insensitive neurons by electrophysiological recordings in PO/AH neurons in slices and study the modulation of their activity by histamine and the ion conductances involved (Specific aim1). The receptor subtypes involved will be studied in Specific Aim2 which will use pharmacological tools, electrophysiology, and single cell RT- PCR. The influence of histamine signaling in the PO/AH on core body temperature and energy expenditure will be investigated in Specific Aim3 which will use telemetry, indirect calorimetry and ex-vivo quantification of the expression of uncoupling proteins 1, 2 and 3 at the mRNA and protein levels. Specific aim 3 will be studied both in w-t and ob/ob mice and the results will be compared in order to determine possible differences in histamine signaling. PUBLIC HEALTH RELEVANCE: Temperature homeostasis is strictly regulated and remarkably stable throughout life and is a key factor to metabolic processes. Histamine signaling in the hypothalamus is a major factor in determining energy homeostasis, thermoregulation, sleep and feeding. These studies will therefore have implications for understanding metabolic disorders, such as obesity and diabetes, as well as sleep disorders such as excessive day sleepiness and narcolepsy.